Although HIV spreads systemically, the majority of infections are transmitted at mucosal surfaces, hence a vaccine should ideally induce protective local immunity. Furthermore, it is now widely felt that potent antiviral cytotoxic T lymphocytes (CTL) should be a major component of any immune response that is induced by an effective vaccine against HIV/AIDS. In this context, it is becoming increasingly apparent that not only the magnitude but also the quality of the T cell response may be important for protection against some pathogens, particularly those proven to be resistant to existing immunization strategies. T cells that show high levels of sensitivity to antigen are said to have 'high functional avidity' and may have the capacity to recognize infected cells very early in infection leading to more efficient recovery or to protection following vaccination. Unfortunately, there are few vaccination strategies currently available that will direct CTL responses to mucosal surfaces, particularly the genital and rectal tissues where the virus is usually first encountered. In this project, we will evaluate genetic vaccination strategies designed to generate mucosal T cell responses, particularly CTL, of high avidity for the immunizing antigen in key mucosal tissues. In particular, we will consecutively deliver intranasal inocula of DNA vaccines and recombinant poxvirus or adenovirus vectors encoding a model vaccine antigen and compare these approaches in terms of their capacity to generate enhanced mucosal T cell responses. The use of ovalbumin (OVA) as our vaccine antigen will facilitate studies of the development, avidity, persistence and migration of vaccine-induced T cell populations due to the variety of analytical reagents that are available in this system. Systemic use of these vectors in this manner resulted in sustained CTL responses of high avidity that expanded rapidly upon re-exposure to recall antigen but these were not expressed in mucosal tissues. It is in these tissues that strong CTL responses, particularly those of high avidity, would be most effective, with the possibility of mucosal elimination of HIV, preventing subsequent systemic infection and establishment of the virus in the host. The capcity of our mucosal vaccination strategy to focus CTL responses in key mucosal tissues may be critical for mucosal containment of the virus and prevention of systemic spread. [unreadable] [unreadable]